Evolution of allostery in the cyclic nucleotide binding module: A comparative genomics study

The cyclic nucleotide binding (CNB) domain regulates signaling pathways in both eukaryotes and prokaryotes. In this study, we analyze the evolutionary information embedded in genomic sequences to explore how the CNB domain elicits a cellular response upon binding to cAMP. Toward this end, we have identified and classified CNB domain sequences in the Global Ocean Sampling data and other protein databases. A systematic comparison of the identifed sequences reveals that the CNB domain recombine with a wide variety of functional domains to acheive functional specificity in signaling pathways. CNB domains have also undergone major sequence variation during evolution. In particular, the sequence motif that anchors the cAMP phosphate (termed the PBC motif) is striking different in some families. This variation may contribute to ligand specificity inasmuch as members of the prokaryotic cooA family, for example, harbor a CNB domain, that contains a non‐canonical PBC motif and that binds a heme ligand in the cAMP binding pocket. Statistical comparison of the functional constraints imposed on the canonical and non‐canonical PBC containing sequences reveals that a key arginine, which coordinates with the cAMP phosphate, has co‐evolved with a glycine in a distal loop that allosterically couples cAMP binding to distal regulatory sites. In conclusion, our analysis suggests that CNB domains have evolved as a scaffold to sense a wide variety of second messenger signals. Based on sequence, structural and biochemical data, we propose a mechanism for allosteric regulation by CNB domains.